The pyrophosphate ion may be represented by the chemical formula P2O72−, and is sometimes referred to by the abbreviation PPi.
PPi is a biologically significant anion which is involved in many cellular processes, such as cellular adenosine triphosphate (ATP) hydrolysis, DNA and RNA polymerizations, and enzymatic reactions. It has been reported that abnormal PPi levels can lead to vascular calcification resulting in severe medical conditions. The selective detection and quantification of PPi can provide insight into cellular processes and abnormalities.
Considerable efforts have been made to develop chemosensors for the optical detection of PPi. One of the early efforts in fluorescent sensing of PPi involved using a polyamine-attached anthracene derivative in 100% aqueous solution. However, poor sensitivity for PPi has hampered reliable fluorescent detection for PPi.
Pyrosequencing is a DNA sequencing technique that is based on a principle sometimes referred to as “sequencing by synthesis.” Pyrosequencing utilizes enzyme-coupled reactions and bioluminescence to monitor the PPi release in real time. The method can be used for single-nucleotide polymorphism (SNP) analysis and tag sequencing (up to 100 bases), as well as for a whole-genome sequencing. The pyrosequencing technique relies on the enzymatic detection of the pyrophosphate that is released during a DNA polymerase chain reaction (PCR). The enzymatic reactions use two enzymes and two reagents (adenosine phosphosulfate (APS) and luciferin)), as shown in the following Scheme 1. The conversion of luciferin to oxyluciferin releases fluorescence at about 562 nanometers (nm). Luciferase controls the “on” and “off” of its bioluminescence, depending on the availability of ATP generated from PPi.

The application requires the probe to specifically recognize the released PPi, in the presence of structurally similar anionic nucleotides. Enzymatic detection of PPi is a complicated scheme, and contributes to the high cost of DNA sequencing.
With the completion of human genome sequencing analysis, the genome-based medicine has come closer to reality. The high cost associated with the DNA sequencing remains to be an obstacle for achieving economically acceptable full-genome analysis in a clinical setting. Significant interest exists in searching for new methods and technologies that can sequence a human genome for $1,000 or less. Low-cost and faster ways to sequence DNA would revolutionize the use of genetic information and the nature of biological and biomedical research.
One significant challenge in the detection of PPi is the need to differentiate between structurally similar anions. Sensing systems for PPi that have improved sensitivity and selectivity, and are more cost effective are needed.